Hedgehog信号通路抑制剂对肝内胆管癌RBE细胞生物学行为的影响

Effect of Hedgehog signaling pathway inhibitor on biological behavior of intrahepatic cholangiocarcinoma cell line RBE

目的:研究Hedgehog(Hh)信号通路特异性抑制剂环杷明(cyclopamine)对人肝内胆管癌细胞株RBE生物学行为的影响。方法:用台盼蓝染色计数法和MTT比色法检测环杷明对RBE细胞增殖的影响;流式细胞术检测凋亡率,Transwell检测环杷明处理前后RBE细胞侵袭能力的变化,Western blot检测环杷明处理前后RBE细胞中Gli1和MMP-9的蛋白表达变化。结果:环杷明对RBE细胞株增殖的抑制作用呈剂量和时间依赖性。环杷明作用细胞24 h、48 h、72 h后,RBE细胞凋亡率逐渐升高,明显高于对照组的凋亡率。Transwell检测对照组穿透细胞数为154.52±13.61,而实验组穿透数为62.00±12.17,侵袭能力明显下降(P<0.01)。Gli1和MMP-9蛋白均在RBE细胞中表达,环杷明下调RBE细胞的Gli1和MMP-9表达。结论:阻断Hh信号通路能抑制RBE细胞的增殖,促进其凋亡,并抑制其侵袭能力。

AIM：To investigate the effect of cyclopamine,a Hedgehog（ Hh） signaling pathway inhibitor,on the biological behavior of intrahepatic cholangiocarcinoma cell line RBE.METHODS：The proliferation of RBE cells was detected by cell counting with Typan blue staining and MTT assay,and the apoptosis was analyzed by the flow cytometry.The Transwell invasive cabin assay was used to detect the invasion ability,and Western blot was used to determine the protein expression of Gli1 and MMP-9 in the RBE cells before and after cyclopamine treatment.RESULTS：Cyclopamine inhibited the growth of RBE cells in a time-and dose-dependent manner.After cyclopamine treatment for 24 h,48 h and 72 h,the apoptotic rates were significantly higher than those in control group.In control group,the number of cells invading through the Matrigel of invasion chamber was 154.52 ± 13.61,while in experimental group it was 62.00 ± 12.17,indicating that the invasion ability of the cells declined significantly.Furthermore,Western blot showed that the protein levels of Glil and MMP-9 in the RBE cells were decreased after treatment with cyclopamine for 24 h and 48 h.CONCLUSION：Blockage of the Hh signaling pathway with cyclopamine suppresses the proliferation,promotes the apoptosis and inhibits the invasion ability of RBE cells.